An enrichment medium for increasing a very small number of vibrio parahaemolyticus cells to the detection limit of the loop-mediated isothermal amplification (LAMP) assay

We developed an enrichment medium for use with the loop-mediated isothermal amplification (LAMP) assay (enrichment media + LAMP assay) to quickly increase a very small number of Vibrio parahaemolyticus cells to the detection limit of the assay. Thirty-nine different enrichment media were prepared based on evaluating 12 potential ingredients. From our assessment of the 39 media, enrichment medium #36, which contained 2% sodium chloride, 1% proteose peptone no. 2, 0.1% trehalose, 0.5% α-ketoglutaric acid, 0.25% pyruvic acid, and 0.5% yeast extract (pH 8.6), was found to be most effective at enhancing the proliferation of V. parahaemolyticus during incubation for 3 h at 40oC. We compared the detection limits of the LAMP assay, the enrichment medium #36 + LAMP assay, and the cultivation method using bacterial cell and spiked shrimp sample tests. The detection limits of the LAMP assay, the medium #36 + LAMP assay, and the cultivation method were 10(3), 10(0)-10(-1), and 10(-1) CFU ml(-1), respectively. Enrichment medium #36 promoted a 10(3)- to 10(4)-fold increase in the bacterial population, and the detection limit of the enrichment media + LAMP assay was the same as that of the cultivation method.     

Analysis of Passive Motion of Para- and Retropharyngeal Structures During Swallowing Using Dynamic Magnetic Resonance Imaging

The purpose of this study was to analyze passive motion of the para- and retropharyngeal space (PRS) during swallowing using dynamic magnetic resonance imaging (MRI). We conducted a preliminary study involving 30 healthy volunteers who underwent dynamic MRI. Consecutive MRI axial images were obtained by examining the plane parallel to the hard palate at the level of the anterior inferior corner of C2. Anterior displacement of the posterior pharyngeal wall (PPW) was measured as a motion index of pharyngeal contraction. The displacement and internal angle of the bilateral external and internal carotid arteries (ECA and ICA) and the bilateral centroids of the PRS area, as well as the increase in PRS area, were calculated at rest and at maximum pharyngeal contraction. In most participants, the bilateral ECA, ICA, and centroids were anterointernally displaced by pharyngeal contraction. The normalized ECA displacement (r = 0.64, r ² = 0.41), normalized ICA displacement (r = 0.60, r ² = 0.37), and normalized centroid displacement (r = 0.43, r ² = 0.19) were more than moderately positively correlated with the normalized PPW displacement. The normalized PRS area increase (r = 0.35, r ² = 0.12) was weakly positively correlated with the normalized PPW displacement. These results revealed that PRS area increased as the ECA and ICA were drawn anterointernally via its passive motion by pharyngeal contraction.     

Effects of the Amino Acid Constituents of Microcystin Variants on Cytotoxicity to Primary Cultured Rat Hepatocytes

Microcystins, which are cyclic heptapeptides produced by some cyanobacterial species from algal blooms, strongly inhibit serine/threonine protein phosphatase and are known as hepatotoxins. Microcystins have many structural variations, yet insufficient information is available on the differences in the cytotoxic potentials among the structural variants. In this study, the cytotoxicities of 16 microcystin variants at concentrations of 0.03–10 μg/mL to primary cultured rat hepatocytes were determined by measuring cellular ATP content, and subsequently determined by their 50% inhibitory concentration (IC₅₀). Differences in the amino acid constituents were associated with differences in cytotoxic potential. [d-Asp³, Z-Dhb⁷] microcystin-LR exhibited the strongest cytotoxicity at IC₅₀ of 0.053 μg/mL among the microcystin variants tested. Furthermore, [d-Asp³, Z-Dhb⁷] microcystin-HtyR was also highly cytotoxic. These results suggest that both d-Asp and Z-Dhb residues are important in determining the cytotoxic potential of microcystin variants.     

Cytopathologic characteristics and differential diagnostic considerations of osteolytic myxopapillary ependymoma

Myxopapillary ependymoma (MPE) is a rare variant of conventional ependymoma found predominantly in the sacrococcygeal region in young adults and characterized by its distinct epithelial and stromal components (WHO grade I designation). MPE with extensive osteolysis is extremely uncommon and only up to 40 cases have been documented. A case is presented here in which imprint smears of a sacral tumor in an 18‐year‐old man revealed complex papillary structures, small loose clusters, or cord‐like structures of bland tumor cells embedded in a myxoid or mucinous background. The tumor cells possessed uniformly round nuclei with a smooth nuclear outline, fine granular chromatin, and small nucleoli. Slender cytoplasmic fibrillary processes and occasional intracytoplasmic vacuoles were observed. A cytologic diagnosis of a MPE was suggested and histochemical and immunohistochemical studies were conducted on formalin‐fixed, paraffin‐embedded material. Immunohistochemically, the tumor cells showed diffuse and strong membranous and cytoplasmic staining for cytokeratin AE1/AE3, glial fibrillary protein, and S‐100 protein, but negative for epithelial membrane antigen, pan‐neuroendocrine markers (i.e., NSE, chromogranin A, synaptophysin), or brachyury. The proliferative index with MIB‐1 was around 10%. The diagnosis of osteolytic MPE was confirmed based on cytopathologic, histopathological, immunohistochemical results, radiologic findings, and the location of the tumor. We demonstrated here the cytopathological features of osteolytic MPE with emphasis on differential diagnostic considerations. Diagn. Cytopathol. 2014;42:778–783. © 2013 Wiley Periodicals, Inc.     

Elucidation of the reaction mechanism on dry reforming of methane in an electric field by in situ DRIFTs

With increasing expectations for carbon neutrality, dry reforming is anticipated for direct conversion of methane and carbon dioxide: the main components of biogas. We have found that dry reforming of methane in an electric field using a Pt/CeO₂ catalyst proceeds with sufficient rapidity even at a low temperature of about 473 K. The effect of the electric field (EF) on dry reforming was investigated using kinetic analysis, in situ DRIFTs, XPS, and DFT calculation. In situ DRIFTs and XPS measurements indicated that the amount of carbonate, which is an adsorbed species of CO₂, increased with the application of EF. XPS measurements also confirmed the reduction of CeO₂ by the reaction of surface oxygen and CH₄. The reaction between CH₄ molecules and surface oxygen was promoted at the interface between Pt and CeO₂.

In the dry reforming of methane in an electric field, the reaction between CH₄ molecules and surface oxygen was promoted at the interface between Pt and CeO₂.     

Diagnostic yield of endoscopic retrograde cholangiography and of EUS-guided fine needle aspiration sampling in gallbladder carcinomas

Background

Obtaining histological evidence of gallbladder carcinoma (GBC) is difficult due to its extraductal nature, and pathological confirmation remains challenging. We compared the diagnostic value and safety of endoscopic ultrasound-guided fine-needle aspiration (EUS-FNA) with endoscopic retrograde cholangiography (ERC) in patients with suspected GBC.

Patients

Eighty-three patients with GBC were evaluated. Prior to definitive management, pathological evidence of GBC was obtained through either ERC cytopathologic sampling (n?=?33), EUS-FNA (n?=?24) or both (n?=?26).

Results

Among the 83 patients, 59 (71.0%) with biliary obstruction were sampled using ERC with 47.4% (28/59) sensitivity. In 19 of the remaining 31 cases, EUS-FNA sampling had 100% diagnostic sensitivity. Likewise, 50 (60.2%) of the 83 patients with suspected GBC underwent EUS-FNA of regional lymph nodes or the gallbladder (GB) mass itself with 94.8% sensitivity. The overall diagnostic sensitivity rates of ERC and EUS-FNA were 47.4 and 96%, respectively (P?<?0.001). Post-procedural complications were seen in 6.7% of the ERC group (4/59, all were mild pancreatitis), and in none of the EUS-FNA group (P?=?0.10).

Conclusions

Gallbladder carcinoma sampling using ERC and EUS-FNA should be incorporated into the diagnostic workup of GB lesions as complementary tools, and EUS-FNA should be applied in the setting of failed or not indicated ERC.     

GenoType MTBDRplus assay for molecular detection of rifampin and isoniazid resistance in Mycobacterium tuberculosis strains and clinical samples

The purpose of this study was to evaluate the GenoType MTBDRplus assay (Hain Lifescience GmbH, Nehren, Germany) for its ability to detect resistance to rifampin (RIF) and isoniazid (INH) in Mycobacterium tuberculosis clinical strains and directly in clinical samples. A total of 62 clinical strains characterized with the Bactec 460TB system were included. For the INH-resistant strains, the MIC was measured and sequencing was performed. Sixty-five clinical samples from 28 patients (39 smear-positive samples and 26 smear-negative samples) were also tested directly. The corresponding isolates of the clinical specimens were studied with the Bactec 460TB system. The overall rates of concordance of the MTBDRplus assay and the Bactec 460TB system for the detection of RIF and INH susceptibility in clinical strains were 98.3% (61/62) and 79% (49/62), respectively. The rate of concordance between the Bactec 460TB system and the MTBDRplus test for the detection of INH resistance in the group of 27 strains with low-level resistance was 62.9% (17/27), and that for the detection of INH resistance in the group of 21 strains with high-level resistance was 85.71% (18/21). Valid test results were obtained for 78.45% (51/65) of the clinical samples tested. The rates of concordance between both assays for the detection of drug resistance in these samples were 98% (50/51) for RIF and 96.2% (49/51) for INH. Taking into account only one sample per patient, the overall rate of concordance between both tests was 92.85% (26/28). The GenoType MTBDRplus assay is easy to perform and is a useful tool for the management of tuberculosis, as it allows the detection of resistance to RIF and INH in M. tuberculosis strains and also in clinical samples.